Journal of Clinical Medicine Review Current Treatment of Juvenile Myelomonocytic Leukemia Christina Mayerhofer 1 , Charlotte M. Niemeyer 1,2 and Christian Flotho 1,2,* 1 Division of Pediatric Hematology and Oncology, Department of Pediatrics and Adolescent Medicine, Medical Center, Faculty of Medicine, University of Freiburg, 79106 Freiburg, Germany; [email protected] (C.M.); [email protected] (C.M.N.) 2 German Cancer Consortium (DKTK), 79106 Freiburg, Germany * Correspondence: christian.fl[email protected] Abstract: Juvenile myelomonocytic leukemia (JMML) is a rare pediatric leukemia characterized by mutations in five canonical RAS pathway genes. The diagnosis is made by typical clinical and hematological findings associated with a compatible mutation. Although this is sufficient for clinical decision-making in most JMML cases, more in-depth analysis can include DNA methylation class and panel sequencing analysis for secondary mutations. NRAS-initiated JMML is heterogeneous and adequate management ranges from watchful waiting to allogeneic hematopoietic stem cell transplan- tation (HSCT). Upfront azacitidine in KRAS patients can achieve long-term remissions without HSCT; if HSCT is required, a less toxic preparative regimen is recommended. Germline CBL patients often experience spontaneous resolution of the leukemia or exhibit stable mixed chimerism after HSCT. JMML driven by PTPN11 or NF1 is often rapidly progressive, requires swift HSCT and may benefit from pretransplant therapy with azacitidine. Because graft-versus-leukemia alloimmunity is central to cure high risk patients, the immunosuppressive regimen should be discontinued early after HSCT. Keywords: juvenile myelomonocytic leukemia; RAS signaling; hematopoietic stem cell transplantation; 5-azacitidine; myelodysplastic/myeloproliferative disorders; targeted therapy Citation: Mayerhofer, C.; Niemeyer, C.M.; Flotho, C. Current Treatment of Juvenile Myelomonocytic Leukemia. J. Clin. Med. 2021, 10, 3084. https:// doi.org/10.3390/jcm10143084 1. Introduction JMML is a pediatric leukemia with shared features of myelodysplastic and myelo- Academic Editor: proliferative neoplasms, usually manifesting during early childhood with leukocytosis, Rupert Handgretinger thrombocytopenia, pronounced monocytosis, splenomegaly, immature precursors on pe- ripheral blood (PB) smear, and bone marrow (BM) blast count below 20% [1–3]. Its clin- Received: 22 June 2021 ical and hematological picture, as well as natural history and outcome, are remarkably Accepted: 10 July 2021 diverse [4]. The common molecular denominator of JMML is the deregulation of the Published: 13 July 2021 intracellular Ras signal transduction pathway, caused in >90% of cases by mutations in one (or, rarely, more than one) of five primordial genes (PTPN11, NRAS, KRAS, NF1, or CBL)[5]. Publisher’s Note: MDPI stays neutral For most patients, allogeneic hematopoietic stem cell transplantation (HSCT) is the only with regard to jurisdictional claims in curative treatment option, in contrast to a smaller percentage of children who survive published maps and institutional affil- long-term without HSCT and eventually experience spontaneous clinical remissions [6,7]. iations. Clinical and molecular risk factors were established to help predict the disease course and guide therapeutic decisions, including age at diagnosis, percentage of fetal hemoglobin (HbF), platelet count, and aberrant DNA methylation patterns [8,9]. In this article, we review the current knowledge of genetic and epigenetic properties of JMML and provide Copyright: © 2021 by the authors. detailed recommendations for the clinical management of children diagnosed with this Licensee MDPI, Basel, Switzerland. challenging disorder. This article is an open access article distributed under the terms and 2. The Origin of JMML: The Ras Pathway conditions of the Creative Commons The Ras pathway is a sequence of kinases in the cell that serves as a chain of com- Attribution (CC BY) license (https:// munication between extracellular mitogens and the cell nucleus [10]. External cytokine creativecommons.org/licenses/by/ signals, relayed through receptor tyrosine kinases and intracellular adapter proteins, lead 4.0/). J. Clin. Med. 2021, 10, 3084. https://doi.org/10.3390/jcm10143084 https://www.mdpi.com/journal/jcm J. Clin. Med. 2021, 10, 3084 2 of 17 to guanosine exchange factor-mediated transformation of Ras proteins into their active guanosine triphosphate (GTP)-bound state (reviewed in more detail in [11,12]). The Ras signal is terminated by intrinsic Ras phosphatase activity, which converts Ras back to an inactive guanosine diphosphate (GDP)-bound configuration. An additional layer of regulation is provided by GTPase activating proteins (GAPs). Effects of Ras activation include the subsequent phosphorylation of Raf, Mek, and Erk kinases [13–17], activation of the mammalian target of rapamycin (mTOR) axis via phosphoinositide 3-kinase (PI3K) [18], and others [19]. Among nuclear targets are the transcription factors Jun and Fos [20]. Genetic mutations in specific Ras pathway components (PTPN11, NRAS, KRAS, NF1, or CBL), resulting in net hyperactivation of the Ras-GTP-GDP loop, are present in hematopoietic cells of >90% of children diagnosed with JMML [4,21–26]. These can be traced back to early myeloid stem/progenitor cell compartments [27–29], and they are found in patient cord blood samples [24], substantiating their role as initiating events and suggesting the inception of the leukemogenic sequence before birth [30]. Somatic mutations in exons 3 or 13 of the PTPN11 gene are present in ~35% of JMML cases [22,31], resulting in a gain-of-function of the nonreceptor tyrosine phosphatase Shp2 [32]. Somatic mutations in NRAS or KRAS codons 12, 13, or 61, accounting for ~25% of JMML cases [4,25,33], freeze Ras in its active GTP-bound form by inhibition of GTPase activity or resistance to GAPs [4]. Somatic PTPN11, NRAS, and KRAS mutations occur in heterozygous form in JMML, indicating strong cell-transforming capacity already in monoallelic fashion. Two congenital developmental disorders predispose to JMML: NF-1 and CBL syn- drome [26,34–36]. Here, the germline of the patient carries a monoallelic loss-of-function mutation of the NF1 or CBL gene, which may have been inherited or arisen de novo. JMML develops after somatic biallelic inactivation of the respective gene in hematopoietic progenitor cells, predominantly by mitotic gene recombination resulting in uniparental isodisomy [21,37]. NF1 functions as a Ras-GAP and thus negatively regulates the Ras pathway [38,39]. Indicative features in children with JMML/NF-1 are the presence of ≥6 cutaneous café au lait spots and/or the family history; other characteristics of NF-1, such as neurofibromas, optic pathway gliomas, bone lesions and neurological abnormali- ties, usually manifest only later. Overall, 10–15% of JMML cases are driven by NF1 [33,40]. CBL is a E3 ubiquitin ligase mediating the decay of receptor tyrosine kinases in the Ras pathway. Mutations targeting exons 8 or 9account for ~15% of JMML cases [26,33]. CBL syndrome, a Noonan-like rasopathy, has a wide phenotypic spectrum. Features include impaired growth, facial anomalies, developmental delay, cryptorchidism, autoimmune phenomena, and notably, neurovasculitis [26,37]. However, it is not rare for patients with JMML and CBL germline mutation to display no abnormalities at all [26,41,42]. Noonan syndrome (NS), the most common rasopathy with an incidence of 1 in 1000–2500 children [43], bears clinical similarities with Turner syndrome. Patients with NS exhibit a short statue, facial dysmorphism, congenital heart defects, skeletal defects, a webbed neck, mental retardation, and cryptorchidism. The genetic basis is a germline mutation in PTPN11 (around 50% of NS cases), SOS1, RAF1, KRAS, BRAF, NRAS or other members of the RAS pathway [5,22,44,45]. Children with NS may experience a polyclonal myeloproliferative disorder (MPD) at a very young age, sometimes shortly after birth [4,46]. Although the condition is indistinguishable from JMML by clinical and hematological fea- tures, it has a self-limiting course in the vast majority of cases. Only a small fraction of children with NS/MPD progress to JMML, presumably after the acquisition of additional genetic changes [5,47]. Although the landscape of PTPN11 mutations is not identical in JMML and NS/MPD [31], there is considerable overlap, and it is not well understood how the same mutation elicits a transient disorder when present in the germline and a fatal disorder when acquired somatically. Obviously, the occurrence of germline and somatic Ras pathway mutations in the same clinical context requires analysis of non-hematopoietic tissue (e.g., hair follicles or skin fibroblasts) to differentiate these conditions [9]. J. Clin. Med. 2021, 10, 3084 3 of 17 Systematic exome sequencing studies revealed that JMML is generally characterized by a paucity of somatic mutations in the neoplastic clone when compared to most other types of cancer [48]. However, subclonal secondary gene mutations can be found in up to half of the cases [23,24,48]. These mutations primarily target the SETBP1, JAK3, SH2B3, or ASXL1 genes. Not infrequently, the secondary mutations affect the Ras pathway itself (“Ras double mutants”). In addition, a role for subclonal mutations in the Polycomb Repressive Complex 2 network was highlighted [23]. Several studies have linked the presence of secondary mutations